Cementum plays a critical role in anchoring the periodontal ligament to the tooth root. However, at present, the mechanisms involved in cementum formation in humans are poorly understood. The major goal of this proposal is to provide basic information regarding human cementum cells and to start elucidating the basic principles involved in the unique, non-lamellar organization of cementum matrix. Since cementum physiology differs significantly among species, the focus will be on human cementum cells. This laboratory has established a technique to culture non-transformed and non-immortalized human cementum-derived cells (HCDC) and to develop an in vivo model to study differentiation of these cells by transplanting them into immunodeficient mice. A combined in vitro/in vivo approach using a large pool of cellular strains that are capable of forming cementum-like tissue in vivo will be carried out. The following two specific aims are proposed in order to better understand the mechanisms of human cementogenesis, especially those relevant to tissue structure and function. Aim 1 proposes to test the hypothesis that the in vivo cementogenic potential of a given cellular strain is associated with the expression of several mineralized tissue markers, cementum-specific attachment protein (CAP) and two small keratan sulfate proteoglycans during cellular differentiation in vitro. In Aim 2, the overall goal is to start elucidating the basic principles involved in the non-lamellar organization of the cementum matrix. To accomplish this, two relevant hypotheses will be tested. The first hypothesis is that the unique, non-lamellar organization of the organic cementum matrix is associated with specific posttranslational modifications of type I collagen. The second hypothesis is that two small, collagen-binding keratan sulfate proteoglycans, lumican (LM) and fibromodulin (FM), play a critical role in the unique non-lamellar organization of the cementum matrix. Collectively, these studies should provide further insight into human cementum physiology at cellular and biochemical levels. A plan to establish the molecular profile of a committed cementogenic cell and to investigate the basic principles of the unique organization of cementum matrix is presented. It is hoped that these studies will lay a foundation for developing therapies aimed at stimulating human cementum growth and/or regeneration.